Doxoruoicin is an antineopiastic agent used to treat a wide variety of tumors. The long term outcome of aoxoruoicin therapy has been disappointing with many toxic side effects. Targeting of cytotoxic agents selectively to cancer cells reduces the toxicity. Luteinizing Hormone Releasing Hormone (LHRH) is a good carrier for cytotoxic agents because its receptors are overexpressed in many cancers, and appear during the process of carcinogenesis, while it is usually not present in normal tissues, with a few exceptions. Attaching a ligand of these receptors to doxorubicin (AN152), decreases the toxicity of this molecule and selectively targets some cancers. These receptors can be activated by phosphorylation and inactivated by dephosphorylation. Epidermal Growth Factor (EGF) phosphorylates LHRH receptors (LHRHR); and Somatostatin (SS) dephosphorylates LHRHR. Through hormonal regulation of LHRHR expression, some malignant cells could be sensitized to AN152. This ongoing study intends to add ress the following questions concerning this new strategy for chemotherapy: (1) Is AN152 selective for LHRHR positive cells? (2) What is the mechanism of AN152 action? Does it enter the cells and act in the nucleus? (3) Can phosphorylation enhance the entry and cytotoxicity of AN152? These questions are being addressed by cell survival studies and fluorescent imaging. The LD50 will be established for AN152 and pretreatment with EGF or RC-160 (an analog of SS) followed by AN152 treatment tor the MCF-7 cells (LHRHR positive human breast carcinoma cells). To assess the mechanism of action, the process is visualized by attaching the chromophore C625 to AN152, to form the compound AN152:C625. Cells are pretreated with EGF or RC-160 and then treated with AN152:C625, or treated with AN152:C625 alone. The uptake of the drug is then tracked by two-photon confocal microscopy. Control experiments are also performed to help define the answers to the posed questions. Free LHRH competition assays, should demonstrate blocking of AN152 uptake and action. Chromophore labeled LHRH (LHRH:C625) is used to help determine the mechanism of AN152 action. Doxorubicin is used in survival studies and one photon confocal microscopy fluorescent imaging (doxorubicin has autofluorescent properties); doxorubicin should be effective independently of LHRHR expression. LHRHR negative cells (UCI-107 human ovarian carcinoma cells) are used in survival studies and two photon confocal microscopy fluorescent imaging. The development of AN152 targeted chemotherapy and its enhancement by EGF could result in effective treatment of many cancers with very low toxic side effects. Key Words: targeted chemotherapy, cancer, LHRH, EGF, two photon confocal scanning laser microsocopy